Data were collected and analyzed using PClamp (Molecular Devices, Union City, CA) and are expressed as means ± S.E.M. At 60 minutes after HFS or LFS, fEPSPs were normalized to baseline recordings (taken as 100%). A two-tailed Student’s t test was used for all comparisons of fEPSPs between groups. When appropriate (Fig. 1), paired t tests were used. Statistical comparisons were based on analysis of IO curves at baseline and 60 minutes after HFS or LFS to determine the degree of change in the maximal rising slope of fEPSPs evoked by stimuli at the 50% point on the IO curves, with P < 0.05 considered significant (Izumi and Zorumski, 2020). For behavioral studies in Fig. 4, data were analyzed by one-way analysis of variance followed by Dunnett’s multiple comparisons test. Numbers reported in the text for statistical analyses are the number (N) of animals studied in a given condition. Statistics were performed using commercial software [SigmaStat (Systat Software, Inc., Richmond City, CA) or GraphPad Prism version 8.3.0 for Windows (www.graphpad.com; GraphPad Software, San Diego, CA)]. Data in figures showing fEPSPs display continuous monitoring of responses at low frequency and thus may differ from numerical results described in the text, which are based on analyses of IO curves.

24S-HC and a synthetic oxysterol analog prevent effects of ethanol on NMDA EPSPs. (A) The graph shows the time course of change of isolated NMDAR-mediated fEPSPs in the presence of 1 μM 24S-HC (blue bar, denoted as 24S in the graph) followed by 60 mM ethanol (denoted as EtOH in the graph) normalized to baseline (100%). In the absence of 24S-HC, ethanol depressed NMDAR-mediated fEPSPs. (B) Even when administered after 60 mM ethanol, which acutely inhibits NMDAR responses, 24S-HC dampened the block of NMDA fEPSPs. (C) SGE-301 (red bar), a synthetic mimic of 24S-HC, also prevents effects of ethanol on NMDA fEPSPs. (D) Akin to 24S-HC, SGE-301 reverses the effects of ethanol on NMDA fEPSPs when administered after block is established. Traces to the right of the graphs show representative NMDA fEPSPs at the times denoted, with baseline responses shown as dashed lines. Calibration: 1 mV, 5 milliseconds.

SGE-301 overcomes the effects of ethanol on one-trial inhibitory avoidance learning. The top bar depicts the latency for rats to enter the dark chamber in the absence of drug treatment (saline controls) 24 hours after receiving a shock upon entry into the dark compartment during conditioning. All of these animals learned the task and failed to enter the dark chamber during the 300-second test. Rats treated with 2 g/kg ethanol 15 minutes prior to the conditioning trial showed a marked decrement in learning and readily entered the dark chamber. Pretreatment with SGE-301 prevented the acute effects of ethanol on learning compared with controls pretreated with vehicle alone (CDX) prior to ethanol. SGE-301 alone had no effect on learning compared with vehicle controls. ****P ≤ 0.0001 by one-way ANOVA, followed by Dunnett’s multiple comparisons test vs. ethanol (F = 17.60).

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